A regulatory protein initially discovered as an activator of cyclic nucleotide phosphodiesterase (or 3',5'-cyclic nucleotide 5'-PDE, EC Kinetic studies1 on the activation of cyclic nucleotide PDE as a function of cal-modulin and Ca2+ concentrations were analyzed by what is likely to be a general approach for characterizing activation mechanisms. The method accounts for the various interactions between phosphodiesterase and different calmodulin-Ca2+ complexes. All four Ca2+ must be bound to calmodulin for the protein to form an activated complex with PDE. The authors identified several regulatory advantages of engaging the four Ca2+ sites on calmodulin: (1) The activation of PDE as a function of Ca2+ is highly cooperative (if the PDEcalmodulin-(Ca2+)4 complex is the dominant active species), and the system behaves as an ''on/off'' switch for PDE activation. (2) At normal intracellular Ca2+ levels (<0.1 ^M), PDE and calmodulin fail to complex, thereby allowing the distribution of calmodulin complexes with its various target enzymes to be reshuffled after resequestration of Ca2+ following each surge. (3) The affinity between the enzyme and the fully liganded calmodulin is 104-105 times that observed in the absence of Ca2+. Thus, a very sharp increase in affinity is achievable through a 10- to 20-times increase in the affinity of Ca2+ for the enzyme-calmodulin complex in each of the four binding steps.

1C. Y. Huang, V. Chau, P. B. Chock, J. H. Wang & R. K. Sharma (1981) Proc. Natl. Acad. Sci. U.S.A. 78, 871.

Selected entries from Methods in Enzymology [vol, page(s)]: Calmodulin purification and fluorescent labeling, 102, 1; assay of calmodulin by Ca2+-dependent phosphodiesterase, 102, 39; myosin light chain phosphorylation in smooth muscle and nonmuscle cells as a probe of calmodulin function, 102, 62; spectroscopic analyses of calmodulin and its interactions, 102, 82; Ca2+ binding to calmodulin, 102, 135; preparation of fluorescent labeled calmodulins, 102, 148; techniques for measuring the interaction of drugs with calmodulin, 102, 171; detection of calmodulin-binding polypeptides separated in SDS-polyacrylamide gels by a sensitive [125I]calmodulin gel overlay assay, 102, 204; use of calmodulin affinity chromatography for purification of specific calmodulin-depen-dent enzymes, 102, 210; chemical approaches to the calmodulin system, 102, 296; 13C chemical shift, 239, 369; contamination in creatine phosphokinase, 238, 76; removal of endogenous calmodulin, 238, 77; heteronuclear relaxation studies, 239, 564; hydrophobic affinity partitioning, 228, 257; myosin light-chain kinase peptide complex, 239, 664, 682, 708; regulation of adenylyl cyclase, 238, 77; as retroviral protease substrate, 241, 290; 3D ^-detected [13C-1H] long-range correlation spectrum, 239, 97.

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